Werner, Halina
(2018)
STRUCTURAL AND FUNCTIONAL IMPACTS OF CHEMICAL MODIFICATIONS IN PEPTIDES AND SMALL PROTEINS.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
Due to their presence in all forms of life, proteins have remained at the forefront of research in myriad disciplines. Chemists employ covalent modifications to probe protein structure and function. At the simplest level of structure, a naturally occurring protein comprises a linear sequence of L-α-amino acids, each of which displays one of twenty sidechains. Simple primary sequence modifications can impact the complex structure and function of proteins in interesting ways. In this dissertation, we document our efforts to understand this relationship through chemical modification of peptides and small proteins.
The sequences of natural proteins can be partially substituted with unnatural amino acids, generating heterogeneous-backbone foldamers. The structures and functions of bioactive proteins have been recreated by heterogeneous-backbone foldamers. These protein mimics show increased resistance to proteolytic digestion; however, no effort has yet determined the relative protection efficiencies of unnatural residues. Thus, we ranked the proteolytic protection imparted by four commonly utilized unnatural residues substituted into a host peptide. These rankings have since been employed in the design of bioactive protein mimics.
Covalent cross-linking of protein sidechains is one type of modification employed to stabilize protein structures. Cross-links made across α-helices are thought to impart structural stability by preorganizing the backbone into an α-helical conformation; however, there is limited
evidence bearing directly on this hypothesis. Thus, we compared the thermodynamic impacts of three cross-link types on folding in a small α-helix rich protein. Our results support the preorganization hypothesis, showing a decreased entropic penalty of folding for all three cross-linked proteins. This finding should help inform the endeavors of the peptide and protein cross-linking community.
The ability of heterogeneous-backbone foldamers to mimic bioactive proteins highlights their potential use as therapeutics. While our group has been developing design principles, we have not yet achieved structural and functional mimicry of a protein larger than 56 residues. Thus, we use native chemical ligation to generate a library of heterogeneous-backbone ubiquitin foldamers, some of which fold and function comparably to native ubiquitin. In addition to refining our design principles, this is the first example of a heterogeneous-backbone foldamer of this size and functional ability.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
27 September 2018 |
| Date Type: |
Publication |
| Defense Date: |
18 July 2018 |
| Approval Date: |
27 September 2018 |
| Submission Date: |
6 August 2018 |
| Access Restriction: |
No restriction; Release the ETD for access worldwide immediately. |
| Number of Pages: |
175 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
Dietrich School of Arts and Sciences > Chemistry |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
ubiquitin foldamers,
unnatural amino acids,
proteolytic stability,
side chain cross-links,
heterogeneous-backbone foldamers,
protein and peptide backbone modification |
| Date Deposited: |
28 Sep 2018 00:08 |
| Last Modified: |
28 Sep 2018 00:08 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/35096 |
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